RESUMO
The nucH gene, encoding a thermostable nuclease (TNase), was isolated from the cellular DNA of Staphylococcus hyicus strain E80 and sequenced. NucH, the 169-amino-acid (aa) protein encoded by this gene, contains, at its N-terminus, a signal peptide which appears to be cleaved at the same site in S. hyicus and Escherichia coli, yielding a mature protein which is exported extracellularly from S. hyicus, but not from E. coli. The aa sequence of NucH is highly homologous with that of the TNase from S. intermedius strain LRA076, whereas significant similarities are observed with the TNase from S. aureus, as well as with three other bacterial proteins of which only one has been shown to exhibit DNase activity. As seen in a multiple sequence alignment, the invariant residues are mostly located in the regions involved in the biological activity of the S. aureus TNase. The ability of crude cell extracts of E. coli strains carrying nucH to degrade various forms of nucleic acids with or without Ca2+ supplementation was studied. Under our experimental conditions, the enzyme encoded by nucH was active at 37 degrees C on both DNA and RNA, had the potential to act as an endonuclease, and functioned in the presence of Ca2+. Moreover, activity was retained after heating at 100 degrees C, suggesting that the enzyme could undergo reversible unfolding.
Assuntos
Nuclease do Micrococo/genética , Staphylococcus/enzimologia , Sequência de Aminoácidos , Sequência de Bases , DNA Bacteriano , Regulação Bacteriana da Expressão Gênica , Nuclease do Micrococo/química , Nuclease do Micrococo/metabolismo , Dados de Sequência Molecular , Mapeamento por Restrição , Homologia de Sequência de Aminoácidos , Staphylococcus/genéticaRESUMO
The Staphylococcus aureus plasmid gene, vgaB, conferring resistance to streptogramin (SgA) and related compounds (PIIA, virginiamycin M, mikamycin A, synergistin A, Dalfopristin) was cloned and sequenced. This gene potentially encodes a 552-aa protein, VgaB, of 61,327 Da, which exhibits a significant similarity with the ATP-binding domains of numerous proteins. VgaB has two ATP-binding domains containing each of the A and the B motifs described by Walker et al. [Walker, J.E., Saraste, M., Runswick, M.J., Gay, N.J., 1982. Distantly related sequences in the alpha- and beta-subunits of ATP synthase, myosin, kinases and other ATP-requiring enzymes and a common nucleotide binding fold. EMBO J., 1, 945-951], but does not include TM hydrophobic domains. The 155-amino-acid sequence between the two ATP-binding domains of VgaB is richer in Glu than the rest of the protein. The vgaB gene was found in 21 of the 52 SgA(R) and independent wt staphylococci investigated. In each of the 21 staphylococci, vgaB was carried on a plasmid of 50-90 kb also harboring the vatB gene encoding an acetyltransferase inactivating SgA. In all plasmids, vgaB and vatB have the same relative positions.
Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Proteínas de Bactérias/genética , Genes Bacterianos , Staphylococcus aureus/genética , Virginiamicina/farmacologia , Transportadores de Cassetes de Ligação de ATP/isolamento & purificação , Sequência de Aminoácidos , Proteínas de Bactérias/isolamento & purificação , Sequência de Bases , Clonagem Molecular , Resistência Microbiana a Medicamentos , Dados de Sequência Molecular , Fenótipo , Plasmídeos/genética , Mapeamento por Restrição , Análise de Sequência de DNA , Staphylococcus aureus/química , Staphylococcus aureus/efeitos dos fármacosRESUMO
The Staphylococcus aureus plasmids, pIP680 and pIP1156, which confer resistance to A-type compounds of virginiamycin-like antibiotics (Vml: streptogramin A, pristinamycin IIA, virginiamycin M) and to synergistic mixtures of the A and B compounds of Vml antibiotics, were shown to direct the modification of A-type compounds by acetylation. The vat gene, encoding the acetyltransferase modifying A-type compounds, was isolated from plasmid pIP680 and sequenced. This gene potentially encodes a 219-amino-acid (aa) protein, VAT, of 24 330 Da showing at least 38% aa identity with two chloramphenicol acetyltransferases encoded by cat genes isolated from Escherichia coli and Agrobacterium tumefaciens. Resistance to A-type compounds of Vml antibiotics conferred to S. aureus by vat was not expressed in E. coli, although a protein having a M(r) similar to that encoded by this gene was detected in E. coli minicells. The vat gene was detected by the polymerase chain reaction in two chromosomally located staphylococcal conjugative elements and in the conjugative plasmid, pIP1156, conferring resistance to A-type compounds.
Assuntos
Acetiltransferases/genética , Proteínas de Bactérias , Resistência Microbiana a Medicamentos/genética , Genes Bacterianos , Fatores R/genética , Staphylococcus aureus/genética , Virginiamicina/farmacologia , Acetiltransferases/química , Sequência de Aminoácidos , Sequência de Bases , Cloranfenicol O-Acetiltransferase/genética , Clonagem Molecular , Contraindicações , DNA Bacteriano/análise , Dados de Sequência Molecular , Mapeamento por Restrição , Homologia de Sequência de Aminoácidos , Staphylococcus aureus/enzimologiaRESUMO
The recombinant plasmid pIP1713 was constructed to analyse the transpositional activity of the insertion sequence IS1181 in Staphylococcus aureus RN4220, Staphylococcus carnosus TM300 and Listeria monocytogenes EGD. This 11.3-kb plasmid contains two genetically different elements: (i) a pE194ts-derived replicon, the ermC gene of which confers resistance to erythromycin in Gram-positive bacteria of several species, and (ii) a copy of IS1181, cloned from S. aureus BM3121, in which the tetracycline resistance gene, tet(T), has been inserted between the transposase-encoded gene and the downstream inverted repeat. When introduced by electroporation into the three bacterial hosts, pIP1713 delivered IS1181 omega tet(T) to various chromosomal sites. Cointegrate structures between pIP1713 and the host chromosome were occasionally detected. Transposition was associated with 8-bp repeats at the insertion sites. IS1181 omega tet(T) could be used for random mutagenesis in Gram-positive bacteria.
Assuntos
Elementos de DNA Transponíveis , Genoma Bacteriano , Listeria monocytogenes/genética , Mutagênese Insercional/métodos , Staphylococcus aureus/genética , Staphylococcus/genética , Sequência de Bases , Clonagem Molecular/métodos , DNA Bacteriano/química , DNA Bacteriano/genética , Vetores Genéticos , Plasmídeos , Mapeamento por RestriçãoRESUMO
Detection of an unusual combination of four resistance markers among coagulase-negative staphylococci (CNS) isolated in the same intensive care unit led to the undertaking of an epidemiological assessment. Seventeen CNS isolates from the same unit and 38 epidemiologically unrelated Staphylococcus epidermidis isolates were typed by eight methods, including analysis of immunoblot patterns and hybridisation patterns (HP) obtained with three probes. The probes comprised plasmids carrying the genes encoding 16S rRNA (pBA2), aacA-aphD (pSF815A), and aacA-aphD with part of IS256 (pIP1307). Immunoblot patterns and HP with pIP1307 indicated that 14 of the 17 CNS isolates from the same unit resulted from the spread of an epidemic strain.
Assuntos
Coagulase/metabolismo , Unidades de Terapia Intensiva , Staphylococcus/isolamento & purificação , Proteínas de Bactérias/análise , Métodos Epidemiológicos , Humanos , Immunoblotting , Testes de Sensibilidade Microbiana , Paris , Plasmídeos , Sorotipagem , Staphylococcus/enzimologiaRESUMO
During a 14-month period, from December 1984 to February 1986, 630 Staphylococcus aureus isolates were identified at Broussais Hospital. Thirty-eight isolates (6%), from 35 patients, were found to be pefloxacin-resistant S. aureus (PRSA) with minimal inhibitory concentrations greater than or equal to 8 mg l-1. PRSA isolates were tested for susceptibility to 35 antibiotics, including nine quinolones, and heavy metal ions. Phage-type was determined. Out of the 38 PRSA isolates, 35 (92%) were methicillin- and multiply-resistant; however, all PRSA isolates were sensitive to vancomycin and coumermycin. Fifteen isolates (39%) had similar phage-type and identical antibiotic susceptibility pattern with high level resistance to pefloxacin (MICs equal to 64 mg l-1); they were isolated from the same surgical unit. The 23 remaining PRSA isolates differed by their phage and susceptibility patterns. Pefloxacin MICs ranged from 8 to 512 mg l-1 with a bimodal distribution; cross-resistance was observed with the eight other quinolones tested. Only nine PRSA isolates (24%), including four 'epidemic' isolates, were obtained from patients who had been treated with quinolones. From these data there is apparently no direct relationship between quinolone administration and selection of PRSA in infected patients.
Assuntos
Pefloxacina/farmacologia , Staphylococcus aureus/efeitos dos fármacos , Adulto , Idoso , Idoso de 80 Anos ou mais , Tipagem de Bacteriófagos , Resistência Microbiana a Medicamentos , Feminino , Hospitais Universitários , Humanos , Masculino , Meticilina/farmacologia , Resistência a Meticilina , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Estudos Prospectivos , Especificidade da Espécie , Staphylococcus aureus/classificaçãoRESUMO
A gene encoding an acetyltransferase inactivating streptogramin A (SgA) and structurally similar compounds was isolated from a staphylococcal plasmid and sequenced. This gene, designated vatB, potentially encodes a 212-amino-acid protein, VatB, of 23,320 Da with 47.4 and 58.4% amino acid identities with two other enzymes with the same activity, Vat and SatA, respectively, which are encoded by a staphylococcal plasmid and an enterococcal plasmid, respectively. The C-terminal parts of these three enzymes share significant homology with the C-terminal parts of 10 other acetyltransferases modifying various substrates. A pair of degenerate primers representing the conserved motifs shared by VatB, Vat, and SatA was designed to detect the three genes encoding these SgA acetyltransferases. Five of 12 clinical SgAr Staphylococcus aureus isolates tested carried neither these genes nor the gene vga, which confers resistance to SgA by a different mechanism, suggesting that another gene(s) and possibly another mechanism of resistance to SgA in staphylococci remains to be characterized.
Assuntos
Acetiltransferases/metabolismo , Antibacterianos/metabolismo , Bactérias Gram-Positivas/enzimologia , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/genética , Virginiamicina/metabolismo , Acetiltransferases/genética , Sequência de Aminoácidos , Antibacterianos/farmacologia , Sequência de Bases , Conjugação Genética , Meios de Cultura , DNA Bacteriano/análise , DNA Bacteriano/isolamento & purificação , DNA Bacteriano/metabolismo , Genes Bacterianos , Bactérias Gram-Positivas/efeitos dos fármacos , Immunoblotting , Testes de Sensibilidade Microbiana , Dados de Sequência Molecular , Hibridização de Ácido Nucleico , Plasmídeos , Reação em Cadeia da Polimerase , Virginiamicina/farmacologiaRESUMO
Several staphylococcal plasmids (26-45 kb) carry all three streptogramin-resistance (Sg(R)) genes, vat, vgb, and vga. Seven such plasmids harbored by independent strains belonging to three taxa (Staphylococcus aureus, S. simulans, and S. cohnii subsp. urealyticum) were compared and the deleted derivative of one of them, pIP680 (11.3 kb), carrying the three streptogramin-resistance genes was sequenced. The seven native plasmids had in common a 12.1-kb part cocarrying the three Sg(R) genes. Sequence analysis of pIP680 revealed that the simultaneous presence of these three genes has probably resulted from cointegration of two plasmids: (i) a pAMbeta1-like plasmid harboring vat-vgb and whose replication gene has been inactivated by an IS257 insertion and (ii) a functional vga plasmid whose replication is similar to that of two staphylococcal plasmids, pSX267 and pSK41.
Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Acetiltransferases/genética , Proteínas de Bactérias/genética , DNA Bacteriano , Hemeproteínas/genética , Plasmídeos , Staphylococcus/genética , Virginiamicina/farmacologia , Sequência de Bases , Resistência Microbiana a Medicamentos , Genes Bacterianos , Dados de Sequência Molecular , Staphylococcus/efeitos dos fármacosRESUMO
Streptogramin and related antibiotics are mixtures of two compounds, A and B (e.g. Dalfopristin and Quinupristin), particularly against Gram-positive bacteria. Staphylococci resistant to these mixtures are always resistant to the A compounds but are not necessarily resistant to the B compounds. Resistance to A compounds and to the mixtures is conferred by acetyltransferases or ATP-binding proteins via unknown mechanisms. Several genes encoding each of the two categories of protein have been characterized and regularly detected on plasmids. Genes encoding lactonases, which inactivate B compounds, have been occasionally detected on these plasmids. Staphylococci which harbour plasmids conferring resistance to A compounds should not be treated with the mixtures even if they appear susceptible in vitro. Indeed, susceptibility to the mixtures of staphylococci carrying resistance to A compounds has often been attributed to partial loss of the plasmids conferring this resistance. When staphylococci are constitutively resistant to B compounds, the in vitro activities of the mixtures should be evaluated, because they are better correlated than MICs with their efficacy in therapy.
RESUMO
The nucleotide sequence of a 1883 bp fragment isolated from a resistance plasmid harbored by a Staphylococcus aureus clinical isolate and carrying the gene, vgb, encoding a hydrolase inactivating the B components of virginiamycin family has been determined. The sequence contains one open reading frame which extends from the ATG codon at nt 641 to a TGA codon at nt 1537 and which potentially codes for a protein of 33.035 Da. This value is in agreement with the apparent size (33 kDa) of the protein observed, in minicell extracts. Inactivation of the B components of the virginiamycin antibiotics as well as resistance to these antibiotics were expressed in a virginiamycin sensitive mutant of Escherichia coli recipient containing the gene on a high copy number plasmid.
Assuntos
Genes Bacterianos , Genes , Hidrolases/genética , Fatores R , Staphylococcus aureus/genética , Sequência de Aminoácidos , Antibacterianos/metabolismo , Sequência de Bases , Dados de Sequência Molecular , Mapeamento por Restrição , Staphylococcus aureus/enzimologia , Virginiamicina/metabolismoRESUMO
Most of the aminoglycoside resistant Staphylococcus aureus strains isolated in France are resistant to all the antibiotics belonging to this family. Two aminoglycoside-modifying enzymes were detected in the wild-type strains studied: an APH3'III and an AAC6'-APH2". These strains also carry two types of streptomycin resistance: high-level resistance due to chromosomal mutation(s) affecting ribosome affinity and low-level resistance, the mechanism of which was not characterized. All the aminoglycoside resistance genes were located on the chromosome. DNA fragments of 1.5 and 1.95 kb carrying the aphA and aacA genes, respectively, were isolated, by cloning, from the cellular DNA of a clinical isolate. When these genes were introduced into Escherichia coli and Bacillus subtilis strains, the enzymes synthesized were indistinguishable from those produced by the S. aureus strains. When the cellular DNAs of wild-type and resistant strains were hybridized with the cloned fragments, sequences homologous to the fragment carrying the aphA gene were found to be located at the same chromosomal site, while those hybridizing with the fragment carrying the aacA gene were at different chromosomal sites.
Assuntos
Genes Bacterianos , Staphylococcus aureus/genética , Aminoglicosídeos/farmacologia , Antibacterianos/farmacologia , Mapeamento Cromossômico , Clonagem Molecular , Enzimas de Restrição do DNA , DNA Bacteriano/genética , Desoxirribonuclease HindIII , Resistência Microbiana a Medicamentos , Humanos , Hibridização de Ácido Nucleico , Fatores R , Homologia de Sequência do Ácido Nucleico , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/isolamento & purificação , Transdução GenéticaRESUMO
DNA fragments, 450 bp in length, were amplified by polymerase chain reaction (PCR) from the thermonuclease gene (nuc) carried by seven epidemiologically independent Staphylococcus aureus isolates. Sequencing of the PCR products led us to characterize 210 bp strictly conserved. A 186 bp piece from within this conserved region was cloned into pUC18. The resulting recombinant plasmid, pIP1608, was used as a probe against the cellular DNA of 360 staphylococcal isolates belonging to 28 species. Only the 146 S. aureus isolates, including four which were not thermonuclease producers, had DNA that hybridized with pIP1608. Among the 214 non-S. aureus staphylococci, 55 exhibited a thermonuclease activity. For 32 of these, the enzymatic activity was inhibited by a commercially available polyclonal antiserum directed against the thermonuclease of an S. aureus strain. These results are in favour of the use of pIP1608 as a probe to specifically recognize S. aureus. Furthermore, we propose a method based on colony blot hybridization and potentially useful to enumerate S. aureus cells in biological samples.
Assuntos
Sondas de DNA , Genes Bacterianos , Nuclease do Micrococo/genética , Staphylococcus aureus/isolamento & purificação , Sequência de Bases , Clonagem Molecular , Primers do DNA , DNA Bacteriano/genética , Fluoresceína , Fluoresceínas , Dados de Sequência Molecular , Hibridização de Ácido Nucleico , Análise de Sequência de DNA , Especificidade da Espécie , Staphylococcus aureus/enzimologia , Staphylococcus aureus/genéticaRESUMO
The repeated nucleotide sequence isolated from a methicillin-resistant Staphylococcus aureus isolate displays the characteristic features of an insertion sequence and was named IS1181. It has a size of 1512 bp and consists of a 1359-bp open reading frame that encodes a 439-amino-acid protein which is predicted to be highly basic and 23-bp terminal inverted complementary repeated sequences exhibiting six mismatches. The three copies of IS1181 isolated from distinct parts of the chromosome of S. aureus, BM3121, are flanked at their ends by 8-bp direct repeats, suggesting a duplication of the target sequence. IS1181 exhibits similarities with IS1165 from Leuconostoc mesenteroides and IS1001 from Bordetella parapertusis. IS1181 was detected in at least two to eight copies in 41 of the 52 S. aureus isolates tested, whereas none of the 26 coagulase-negative staphylococci, 24 streptococci, or 11 enterococci analyzed carried nucleotide sequences hybridizing with IS1181.
Assuntos
Elementos de DNA Transponíveis/genética , DNA Bacteriano/química , Nucleotidiltransferases/genética , Staphylococcus aureus/genética , Sequência de Aminoácidos , Sequência de Bases , Primers do DNA , DNA Bacteriano/isolamento & purificação , Proteínas de Ligação a DNA/biossíntese , Proteínas de Ligação a DNA/genética , Escherichia coli/genética , Dados de Sequência Molecular , Nucleotidiltransferases/biossíntese , Fases de Leitura Aberta , Plasmídeos , Reação em Cadeia da Polimerase , Sequências Repetitivas de Ácido Nucleico , Mapeamento por Restrição , Homologia de Sequência de Aminoácidos , Homologia de Sequência do Ácido Nucleico , Staphylococcus aureus/enzimologia , TransposasesRESUMO
EcoRI-digested cellular DNAs of 150 staphylococcal clinical isolates were probed with a plasmid containing a DNA piece from within the open reading frame of IS256. Most of the Gmr staphylococcal isolates tested contained more copies of IS256 than those associated with Tn4001 carried by these isolates. Three distinct copies of IS256 together with their flanking DNA were isolated by cloning from an EcoRI digest of the cellular DNA of the Gmr isolate, BM3121. These three copies of IS256 were shown to be intact. The results from DNA sequencing revealed that one of the three copies is flanked by 8-bp direct repeats, and it is suggested that this may be the result of insertion of the IS256 at this site by autonomous movement of the IS element. The insert DNA of all three clones hybridized to the cellular DNA prepared from a Staphylococcus aureus strain that carries neither IS256 nor the aacA-aphD gene. Two of the clones hybridized to several EcoRI fragments of the cellular DNA of this strain, indicating that in these cases IS256 is flanked by DNA present as several copies on the chromosome.
Assuntos
Elementos de DNA Transponíveis , Família Multigênica , Staphylococcus/genética , Sequência de Bases , Clonagem Molecular , DNA Bacteriano/genética , Humanos , Dados de Sequência Molecular , Hibridização de Ácido Nucleico , Sondas de Oligonucleotídeos , Fases de Leitura Aberta , Plasmídeos , Recombinação Genética , Staphylococcus/isolamento & purificaçãoRESUMO
A variant of the vga(A) gene (1,575 bp), encoding an ATP-binding cassette protein conferring resistance to streptogramin A and related antibiotics, was cloned from the chromosome of a Staphylococcus aureus clinical isolate and sequenced. The sequence of the variant was similar to that of the vga(A) gene (83.2% identity). However, the G+C content of the variant (35.6%) was higher than that of vga(A) (29%) and there was no cross hybridization between vga(A) and the variant at high stringency (> or =60 degrees C), the highest temperature at which a signal was detected being 55 degrees C. Unlike previous reports for vga(A) and vga(B), the variant of vga(A) may be present in multiple copies in the genome. These copies are chromosomal in some isolates and both chromosomal and plasmid-borne in others. Nucleotide sequences hybridizing at 65 degrees C with the vga(A) variant were found in all the staphylococcal strains harboring plasmids carrying both vga(B) and vat(B), which also encode resistance to streptogramin A.
Assuntos
Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Staphylococcus aureus/genética , Virginiamicina/farmacologia , Alelos , Motivos de Aminoácidos , Proteínas de Bactérias/fisiologia , Clonagem Molecular , Primers do DNA , Sondas de DNA , DNA Bacteriano/análise , Resistência Microbiana a Medicamentos/genética , Humanos , Testes de Sensibilidade Microbiana , Dados de Sequência Molecular , Hibridização de Ácido Nucleico , Plasmídeos/genética , Reação em Cadeia da Polimerase , Staphylococcus aureus/efeitos dos fármacos , TemperaturaRESUMO
A new staphylococcal composite transposon, designated Tn5405, carrying the genes aphA-3 and aadE, which encode resistance to aminoglycosides, was partially characterized. The transposon is 12 kb long and is flanked by inverted repeated sequences displaying the characteristic features of an insertion sequence, named IS1182. This insertion sequence is 1864 bp long and has 23/33-bp imperfect inverted repeats at its ends. One of the IS1182 copies delimiting Tn5405 contains a copy of IS1181 flanked by 8-bp direct repeats. Tn5405 was found in the chromosome of MRSA clinical isolate BM3121, within a Tn552-related transposon, Tn5404. Tn5404 was previously characterized following its transposition onto a beta-lactamase plasmid harbored by BM3121. Two forms of the recombinant beta-lactamase-encoding plasmid generated by the inversion of Tn5405 within Tn5404 were detected. IS1182 was not detected in the DNA of 4 of the 17 tested MRSA isolates containing aphA-3 and resistant to streptomycin. Thus, aphA-3 and aadE genes are not disseminated only by Tn5405 or related transposons delimited by IS1182.
Assuntos
Aminoglicosídeos/farmacologia , Elementos de DNA Transponíveis , DNA Bacteriano , Resistência Microbiana a Medicamentos/genética , Staphylococcus aureus/genética , Sequência de Aminoácidos , Sequência de Bases , Inversão Cromossômica , Cromossomos Bacterianos , Dados de Sequência Molecular , Plasmídeos , Mapeamento por Restrição , Análise de Sequência de DNARESUMO
The incidence of infections with phage type 77 methicillin-resistant Staphylococcus aureus (MRSA) strains increased in France in 1987. These strains are widespread in numerous European hospitals. The SmaI restriction profiles of total DNA extracted from 74 phage type 77 MRSA strains isolated from 1987 to 1994 in 10 hospitals in eight European cities (in France, Belgium, and Spain) were analyzed. Hybridization with a probe containing a 468-bp DNA fragment from within the transposase gene of the insertion sequence IS256 was also examined. Forty-three SmaI profiles were detected. Twenty major genotypes were identified, and each genotype contained strains with the same profile or profiles which differed by no more than three bands. Strains isolated in different countries and at several-year intervals were often grouped within the same genotype. A larger number of genotypes could be discriminated by analysis of the patterns of hybridization with the IS256 probe. SmaI restriction fragments with the same apparent electrophoretic mobility could, in some cases, be distinguished by the presence or the absence of nucleotide sequences hybridizing with IS256. The strains that grouped within the same genotype after hybridization with IS256 were mostly those isolated in the same hospital and at less than 12-month intervals. Consequently, the IS256 probe that we used improved restriction profile analysis for discrimination between the intrahospital, outbreak-related phage type 77 MRSA strains and the endemic strains disseminated in various cities and countries.
Assuntos
Técnicas de Tipagem Bacteriana , DNA Bacteriano/análise , Polimorfismo de Fragmento de Restrição , Staphylococcus aureus/classificação , Tipagem de Bacteriófagos , Análise por Conglomerados , Infecção Hospitalar/microbiologia , Sondas de DNA , Elementos de DNA Transponíveis , Desoxirribonucleases de Sítio Específico do Tipo II , Europa (Continente) , Genótipo , Humanos , Resistência a Meticilina/genética , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/genéticaRESUMO
Tn5405 (12 kb) is a staphylococcal composite transposon delimited by two inverted copies of IS1182, one of which contains IS1181. The internal part of this transposon carries three antibiotic resistance genes, aphA-3, aadE, and sat4, and three open reading frames (ORFs), orfx, orfy, and orfz, of unknown function. The dispersion of Tn5405 and the genes and ORFs included in this transposon were investigated in 50 epidemiologically unrelated staphylococci carrying aphA-3. Twenty-three maps, distinguishable by the presence or absence of the investigated genes or ORFs and/or by the sizes of the restriction fragments carrying them, were identified. Four isolates carried Tn5405, and 15 other isolates contained a Tn5405-related element. IS1182 was not detected in the aphA-3 regions mapped in 31 isolates which carried the following combinations: orfx, orfy, aadE, sat4, and aphA-3 +/- orfz; orfy, aadE, sat4, and aphA-3 +/- orfz; and aadE, sat4, aphA-3, and orfz. In all isolates, the genes and ORFs investigated were in relative positions similar to those in Tn5405. Thus, the internal part of Tn5405 appeared to be partially conserved with the maintenance, in all of the isolates, of at least the three antibiotic resistance genes.
Assuntos
Genoma Bacteriano , Staphylococcus/genética , DNA Bacteriano/isolamento & purificação , Resistência Microbiana a Medicamentos/genética , Mapeamento de Nucleotídeos , Reação em Cadeia da Polimerase , Staphylococcus/efeitos dos fármacosRESUMO
Tn5405 is a 12 kb staphylococcal composite transposon delimited by two inverted copies of the insertion sequence IS1182. This transposon carries two aminoglycosides resistance genes, aphA-3 and aadE, an altered gene similar to sat4 from Campylobacter coli BE/G4, and three open reading frames of unknown functions.
Assuntos
Antibacterianos/farmacologia , Elementos de DNA Transponíveis , Resistência Microbiana a Medicamentos/genética , Staphylococcus aureus/genética , Sequência de Aminoácidos , Aminoglicosídeos , Sequência de Bases , Dados de Sequência MolecularRESUMO
A total of 101 staphylococcal strains were ribotyped using EcoRI and HindIII as restriction enzymes and plasmid pBA2 as the rDNA probe. Isolates from 10 newly described staphylococcal taxa were among those examined. All the ribotypes were added to our database, Staph DB, which now contains the sizes of the bands of 135 EcoRI and 120 HindIII ribotypes from 408 strains belonging to 42 staphylococcal taxa. The relatedness of ribotypes was evaluated by using the Dice coefficient. The ribotypes, and thus the strains, were clustered by the unweighted pair group method with averages (UPGMA). Separation into clusters correlated well with the delineation of the staphylococcal species but not with that of the different subspecies. No discrimination was possible between Staphylococcus vitulinus and Staphylococcus pulvereri. Ecovar-specific groups were evident within Staphylococcus intermedius and Staphylococcus hyicus. The data increase the usefulness of rRNA gene restriction site polymorphism analysis for staphylococcal taxonomy.